Direct characterization of the Na+/H+ exchanger in human platelets

The kinetic properties of the Na+/H+ exchanger in human platelets were investigated by direct measurements of pHi as detected with the fluorescent dye, BCECF. In acid-loaded cells, the antiporter displayed a hyperbolic dependence regarding external Na+ with an apparent Km of 38 +/- 4 mM (pHo 7.2 at 25 degrees C) whereas its pHi-dependent activation between 7.3 to 6.4 did not obey a Michaelian model. External acidification from 7.7 to 6.5 decreased significantly the initial rate of Na(+)-dependent H+ efflux. The amiloride derivative, ethylisopropylamiloride blocked this exchanger and exerted a non-competitive inhibition with respect to Na+o (Ki = 17 nM). The cation selectivity of the external site of the antiporter was Na+ greater than Li+ greater than K+ and choline. These results indicate that the BCECF technique allows to evaluate the main features of the Na+/H+ exchanger in human platelets, which possesses kinetic properties similar to those reported in other cell types.